Alternative arrangements of catalytic residues at the active sites of restriction enzymes

FEBS Letters

Tamulaitis, Gintautas²  Solonin, Alexander S.¹  Siksnys, Virginijus² 

[1]Institute of Biochemistry and Physiology of Microorganisms, Russian Academy of Sciences, Pushchino, Moscow Region 142290, Russia

[2]Institute of Biotechnology, Graiciuno 8, Vilnius 2028, Lithuania

关键词: Restriction endonuclease Ecl18kI;    Active site;    Mutagenesis;    aa;    amino acids;    Ap and Apr;    ampicillin and ampicillin resistance;    BSA;    bovine serum albumin;    Cm and Cmr;    chloramphenicol and chloramphenicol resistance;    EDTA;    ethylenediaminetetraacetic acid;    Tris;    tris(hydroxymethyl)aminomethane;    wt;    wild-type;   

DOI  :  10.1016/S0014-5793(02)02621-2

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

A catalytic sequence motif PDX_10–30(E/D)XK is found in many restriction enzymes. On the basis of sequence similarities and mapping of the conserved residues to the crystal structure of NgoMIV we suggest that residues D160, K182, R186, R188 and E195 contribute to the catalytic/DNA binding site of the Ecl18kI restriction endonuclease. Mutational analysis confirms the functional significance of the conserved residues of Ecl18kI. Therefore, we conclude that the active site motif ¹⁵⁹VDX₂₁KX₁₂E of Ecl18kI differs from the canonical PDX_10–30(E/D)XK motif characteristic for most of the restriction enzymes. Moreover, we propose that two subfamilies of endonucleases Ecl18kI/PspGI/EcoRII and Cfr10I/Bse634I/NgoMIV, specific, respectively, for CCNGG/CCWGG and RCCGGY/GCCGGC sites, share conserved active site architecture and DNA binding elements.

【 授权许可】

Unknown